Purging of mammary carcinoma cells during ex vivo culture of CD34+ hematopoietic progenitor cells with recombinant immunotoxins.
Tumor cells have been found in autologous hematopoietic cell transplants used after high-dose chemotherapy. To specifically eliminate contaminating mammary tumor cells during ex vivo expansion of CD34+ hematopoietic progenitor cells, we used recombinant immunotoxins (ITs) directed against cell-surface antigens expressed on mammary carcinoma cells. ITs were expressed from fusion cDNAs combining a single-chain antibody fragment (scFv) directed against the Erb-B2 or epidermal growth factor (EGF) receptors with a truncated Pseudomonas exotoxin A fragment devoid of its cell-binding domain. CD34+ hematopoietic progenitor cells did not express Erb-B2 and EGF receptors as detected by Western blotting. Ex vivo expansion of total hematopoietic cells or of colony-forming cells from CD34+ progenitors in the presence of stem-cell factor (SCF), interleukin-1 (IL-1), IL-3, IL-6, and erythropoietin (Epo) was not affected when ITs were added to the cultures. In contrast, MDA-MB 453 and MCF-7 mammary carcinoma cells were depleted in a dose- and time-dependent manner by more than 3 log in coculture with CD34+ cells over a period of 7 days in the presence of 100 to 1,000 ng/mL of anti-Erb-B2 IT. This included elimination of the subpopulations with regrowth potential. Similarly, addition of either anti-Erb-B2 or anti-EGF receptor ITs to primary breast cancer cells isolated from patients with metastatic disease resulted in elimination of cytokeratin-positive cells in seven of seven samples. ITs are highly efficient and convenient to use for the depletion of mammary tumor cells during ex vivo expansion of hematopoietic progenitor-cell autografts.
Spyridonidis
A
A
Department of Hematology/Oncology, University Medical Center, Freiburg, Germany.
Schmidt
M
M
Bernhardt
W
W
Papadimitriou
A
A
Azemar
M
M
Wels
W
W
Groner
B
B
Henschler
R
R
D036002:ADP Ribose Transferases; D018952:Antigens, CD34 / Q000032:analysis*; D001427:Bacterial Toxins; D001943:Breast Neoplasms / Q000737:chemistry / Q000473:pathology*; D016923:Cell Death; D002469:Cell Separation; D066246:ErbB Receptors / Q000032:analysis / Q000276:immunology; D005098:Exotoxins / Q000008:administration & dosage / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000737:chemistry / Q000473:pathology*; D006801:Humans; D007128:Immunoglobulin Fragments; D000922:Immunotoxins / Q000494:pharmacology*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000032:analysis / Q000276:immunology; D011993:Recombinant Fusion Proteins; D014407:Tumor Cells, Cultured; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D018952:Antigens, CD34; D001427:Bacterial Toxins; D066246:ErbB Receptors; D005098:Exotoxins; D007128:Immunoglobulin Fragments; D000922:Immunotoxins; D018719:Erb-b2 Receptor Tyrosine Kinases; D011993:Recombinant Fusion Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A